Zetasizer nano analyzer

The powder of Coptidis Rhizoma extract was dissolved in water. After ultrasonic treatment for 1 h, the solution was centrifuged at 3000 rpm for 10 min. The obtained supernatant was filtered through a 0.22-μm filter. The filtered solution was then dialyzed in a dialysis bag (3500 D) against water for five consecutive days. Nnps were obtained by freeze-drying the residues in the dialysis bag. In addition, the Nnps and BBR were dissolved in water at a weight ratio of 1:1. The solution was then boiled for 1 h and then lyophilized to obtain the Nnps-BBR complex powder.
The particle size and Zeta potential in the aqueous solution of the Nnps or the Nnps-BBR complex were determined using a Malvern Zetasizer Nano analyzer (Worcestershire, UK). The powder of the Nnps or the Nnps-BBR complex was sprayed with gold, dried in vacuum, and then observed under an FEI Quanta 250 scanning electron microscope (SEM) (Oregon, USA) operating at 10 kV. A Leica SP8 laser confocal fluorescence microscope (LCFM) (Wetzlar, Germany) was used to observe the morphology of the powder of the Nnps or the Nnps-BBR complex. The protein content in the Nnps was determined using a BCA kit. The content of polysaccharide in the Nnps was determined by phenol sulfuric acid method using glucose as a reference standard. The content of alkaloids in the Nnps or the Nnps-BBR complex was determined using the LC-MS method.

To obtain the undenatured Nnps-forming proteins, a new Coptidis Rhizoma extract was prepared according to the method described in section “Preparation and quality control of Coptidis Rhizoma extract”, but the extraction temperature was maintained at 50°C. The proteins in the extract were obtained by dialysis according to the method described in section “Preparation and characterization of Nnps and Nnps-BBR complex”. After dissolving, the purity and molecular weight of the proteins were determined by standard gel electrophoresis (SDS-PAGE) in 4% concentrating gel and 10% separating gel and stained with silver, with a protein gel electrophoresis device (Bio-Rad, USA). In addition, after the protein solution was boiled for 1 h, the particle size and Zeta potential of the formed particles were measured using the Malvern Zetasizer Nano analyzer.